False-positive results obtained from the branch-site test of positive selection

Natural selection operating at the amino acid sequence level can be detected by comparing the rates of synonymous (r(S)) and nonsynonymous (r(N)) nucleotide substitutions, where r(N)/r(S) (omega) > 1 and omega < 1 suggest positive and negative selection, respectively. The branch-site test has been developed for detecting positive selection operating at a group of amino acid sites for a pre-specified (foreground) branch of a phylogenetic tree by taking into account the heterogeneity of omega among sites and branches. Here the performance of the branch-site test was examined by computer simulation, with special reference to the false-positive rate when the divergence of the sequences analyzed was small. The false-positive rate was found to inflate when the assumptions made on the omega values for the foreground and other (background) branches in the branch-site test were violated. In addition, under a similar condition, false-positive results were often obtained even when Bonferroni correction was conducted and the false-discovery rate was controlled in a large-scale analysis. False-positive results were also obtained even when the number of nonsynonymous substitutions for the foreground branch was smaller than the minimum value required for detecting positive selection. The existence of a codon site with a possibility of occurrence of multiple nonsynonymous substitutions for the foreground branch often caused the branch-site test to falsely identify positive selection. In the re-analysis of orthologous trios of protein-coding genes from humans, chimpanzees, and macaques, most of the genes previously identified to be positively selected for the human or chimpanzee branch by the branch-site test contained such a codon site, suggesting a possibility that a significant fraction of these genes are false-positives.     

Canadian experience in cytology proficiency testing

The mandatory Laboratory Proficiency Testing Program (LPTP) of the province of Ontario as applied to cytology laboratories is briefly described. LPTP evaluates reporting of test slides to identify laboratories that have deficiencies. Such laboratories receive an on-site visit, followed by recommendations on means of improvement (especially educational) and subsequent monitoring. Most of the 18 cytology laboratories visited to date have shown an improved level of performance on later tests.     

Scientific issues related to the cytology proficiency testing regulations

The member organizations of the CETC feel strongly that there are significant flaws associated with the proposed proficiency test and its implementation. The most immediate modifications include lengthening the required testing interval, utilizing stringently validated and continuously monitored slides, changing the grading scheme and changing the focus of the test from individuals to laboratory level testing, as described above. Integration of new computer-assisted and location-guided screening technologies into the testing protocol is necessary for the testing program to be compliant with the current CLIA law. The regulation also needs to be flexible enough to accommodate new technologies that are implemented in laboratory practice, education and administration of the test. The changes recommended in this document address the most immediate technical and scientific concerns with the current implementation of PT for gynecologic cytology. The CETC will be submitting a subsequent document, following full review of the current regulations, with recommendations for changes, justifications and impact.     

Comparative study of the ThinPrep Pap test and conventional cytology results in a Canadian cohort

OBJECTIVE: To compare the frequency of Pap test results in a prospective series of direct to vial ThinPrep tests to a cohort of conventionally prepared tests. To follow-up all test results for a minimum of 2 years and assess performance based on this outcome. METHODS: All women presenting for either routine screening or colposcopic examination in 2001 were enrolled in the ThinPrep cohort. A similar, population of conventionally prepared tests was extracted from the year 2000 laboratory data. Information on all concurrent and follow-up cervical specimens over the ensuing 2 years was retrieved. RESULTS: The ThinPrep cohort comprised 2288 Pap tests and the conventional, 2211. The frequency of normal [within normal limits (WNL) and benign cellular changes (BCC)] results in the ThinPrep cohort was 6% lower and the frequency of abnormal [> or =atypical squamous cells of undetermined significance (ASCUS)] results was 6.8% higher. Respective ThinPrep and conventional cohort results were 1156 (51%) and 1291 (58%) WNL, 625 (27%) and 561 (25%) BCC, 101 (4%) and 65 (3%) ASCUS, 21 (1%) and 2 (0.1%) atypical glandular cells of undetermined significance, 301 (13%) and 224 (10%) low-grade squamous intraepithelial lesion (LSIL), and 74 (3%) and 40 (2%) high-grade SIL (HSIL) (P < 0.0001). Follow-up was available for nearly 80% of each cohort. LSIL or higher was confirmed in 57.5% (n = 266) of the abnormal ThinPrep and 60.9% (n = 190) of the abnormal conventional tests. The ThinPrep yield of confirmed tests however was almost 50% higher than the conventional test. CONCLUSION: In this population, ThinPrep was superior to the conventional Pap test.     

False-positive human immunodeficiency virus enzyme immunoassay results in pregnant women

Objective

Examine whether false-positive HIV enzyme immunoassay (EIA) test results occur more frequently among pregnant women than among women who are not pregnant and men (others).

Design

To obtain a large number of pregnant women and others tested for HIV, we identified specimens tested at a national laboratory using Genetic Systems HIV-1/HIV-2 Plus O EIA from July 2007 to June 2008.

Methods

Specimens with EIA repeatedly reactive and Western blot-negative or indeterminate results were considered EIA false-positive. We compared the false-positive rate among uninfected pregnant women and others, adjusting for HIV prevalence. Among all reactive EIAs, we evaluated the proportion of false-positives, positive predictive value (PPV), and Western blot bands among indeterminates, by pregnancy status.

Results

HIV prevalence was 0.06% among 921,438 pregnant women and 1.34% among 1,103,961 others. The false-positive rate was lower for pregnant women than others (0.14% vs. 0.21%, odds ratio 0.65 [95% confidence interval 0.61, 0.70]). Pregnant women with reactive EIAs were more likely than others (p<0.01) to have Western blot-negative (52.9% vs. 9.8%) and indeterminate results (17.0% vs. 3.7%) and lower PPV (30% vs. 87%). The p24 band was detected more often among pregnant women (p<0.01).

Conclusions

False-positive HIV EIA results were rare and occurred less frequently among pregnant women than others. Pregnant women with reactive EIAs were more likely to have negative and indeterminate Western blot results due to lower HIV prevalence and higher p24 reactivity, respectively. Indeterminate results may complicate clinical management during pregnancy. Alternative methods are needed to rule out infection in persons with reactive EIAs from low prevalence populations.     

The frequency of false-positive and false-negative results in the detection of Y-chromosomes in interphase nuclei

Summary In blood smears from 527 females and 457 males examined for the presence of Y chromosomes in interphase nuclei, 0.6% false-positive results and 11% false-negative results were found. There was a clear tendency for the falsenegative results to occur among those with small fluorescent or non-existing bands on the Y chromosome. The three falsepositive females all had fluorescent chromosomal variants. In a comparison between female samples with and without chromosomal variants respectively, the former showed significantly higher false Y-body counts. There was a decrease in the number of Y-bodies with increasing age. There were no significant differences between staining with 0.1% Quinacrine mustard and 0.1% and 1% Mepacrine. This study provides a] more solid basis for the use of Y chromosome detection in forensic medicine, for screening purposes etc.     

False "false-positive" results in diagnostic cytology

With the introduction of transbronchial brushings and fine needle aspiration biopsy, which enable us to obtain samples directly from lesions, the diagnostic potential of cytology for the detection of malignancy, including early cancer, has been greatly enhanced. From 1976 to 1982, five positive cytology reports were initially considered to be "false positives" on the basis of negative gross findings, benign operative biopsies or negative histologic findings in the resected surgical specimens. However, these proved to be false "false positives," based upon the clinical follow-up or further examination of the surgical specimens. Presentation is made of three of these cases with positive cytologic findings and initially negative histologic diagnoses, with an analysis of the causes of the latter. From our experience, four types of cancerous lesions seem prone to being missed during gross examination, namely: any small cancer with a consistency similar to that of the parenchyma of the organ in which the tumor is located, superficially invasive carcinoma, scar cancer and a radiologically occult lung cancer in the presence of a coexisting radiologically demonstrable lesion. With more clinical application of these cytologic methods, false "false positives" are expected to occur more often.     

Reliability of the Amplicor internal control to disclose false-negative Chlamydia trachomatis PCR results

Four possibly false-negative samples were detected when 514 male urine specimens were tested in the Amplicor Chlamydia trachomatis assay. In three of the four samples, the inhibition could be reduced by removal of urine supernatant. Under partially inhibitory conditions, after spiking with 50 C. trachomatis elementary bodies/ml specimen, a selective inhibition of the C. trachomatis target amplification and a preferential internal control amplification was observed. We conclude that a positive internal control signal might be misleading in inhibitory specimens with low amount of C. trachomatis.     

CytoView. A prototype computer image-based Papanicolaou smear proficiency test

OBJECTIVE: To develop and assess CytoView, a prototype computer image-based cytology proficiency testing (PT) system, as an alternative to glass-slide cervical cytology PT. STUDY DESIGN: The computer-based PT consists of 10 cases taken from 10 Pap smears, each of which had received a consensus Clinical Laboratory Improvement Amendments-category diagnosis from three pathologists. Each CytoView "case" was the digital representation of > 8,000 microscopic fields, captured from a selected 5 x 10-mm rectangle of a Pap smear. The 5 x 10-mm capture rectangle was selected from the slide's most representative area for its diagnostic category. The CytoView project team developed each case through a multistep process that included image capture, image alignment, correlation of 10x and 40x images, and image display. The 10-slide CytoView PT prototype was then assessed by groups of cytopathologists and cytotechnologists. RESULTS: The CytoView prototype PT system was developed and assessed. CONCLUSION: Preliminary evaluation of CytoView indicated potential for this format as a valid and logistically feasible alternative to the traditional glass slide PT format.